Hepatitis B virus (HBV) remains an important global pathogen that chronically infects hundreds of millions of people worldwide and causes hepatitis, cirrhosis and liver cancer (Trepo C., et al. (2014). Lancet, 384(9959):2053-2063). HBV is an enveloped virus with an inner capsid shell, which in turn encloses a small (3.2 kbp) DNA genome. As a member of the Hepadnaviridae family, which also includes related animal viruses like the duck hepatitis B virus (DHBV), HBV replicates its DNA genome via reverse transcription of a RNA intermediate called pregenomic RNA (pgRNA) (Summers J, Mason W S (1982). Cell 29: 403-415; Seeger C, et al. (2013) Hepadnaviruses. In: Knipe D M, Howley P M, editors. Fields Virology. Philadelphia: Lippincott, Williams & Wilkins. pp. 2185-2221). HBV assembly begins with the assembly of a nucleocapsid (NC) that packages specifically a copy of pgRNA, in complex with the virally-encoded reverse transcriptase (RT) protein (Hu J, Seeger C (2015) Hepadnavirus Genome Replication and Persistence. In: Seeger C, Locarnini S, editors. Cold Spring Harb Perspect Med: Cold Spring Harbor Laboratory Press). The pgRNA is then converted to DNA within the NC by the RT protein.
The icosahedral HBV capsid shell enclosing the viral RNA or DNA is composed of multiple copies of a single viral protein, the HBV core or capsids protein (C or HBc). This small (ca. 21 kd) protein has multiple essential functions in the viral life cycle: including assembly into capsids, packaging of the viral pgRNA and RT protein, regulation of viral reverse transcription, NC envelopment and virion secretion, and nuclear import of the viral genome (Seeger, C. et al., Id. Hu, J. et al., Id). HBc can be divided into a N-terminal domain (NTD, from position 1-140), responsible for capsid assembly (thus also called assembly domain) and a C-terminal domain (CTD, 149-183 or 185 depending on the strains), which are connected by a linker region (140-149) (Steven A C, et al. (2005). Adv Virus Res 64: 125-164). The basic building blocks of the HBV capsids are HBc dimers, with 90 or 120 dimers self-assembling into T=3 or T=4 icosahedral capsid (Zhou S, et al. (1992). Proc Natl Acad Sci USA 89: 10046-10050; Wynne S A, et al., (1999). Mol Cell 3: 771-780; Wingfield P T, et al., (1995). Biochemistry 34: 4919-4932). In heterologous overexpression systems including bacteria and insect cells and in vitro assembly reactions using purified protein, NTD alone, without CTD, is clearly sufficient for assembly into capsids morphological similar to authentic capsids assembled from full-length HBc (Wingfield P T, et al., (1995). Biochemistry 34: 4919-4932; Lanford R E, et al., (1990). Virology 176:222-233; Birnbaum F, Nassal M (1990). J Virol 64: 3319-3330; Gallina A, et al. (1989). J. Virol 63: 4645-4652). On the other hand, the arginine-rich (protamine-like) and highly basic CTD displays non-specific RNA and DNA binding and nucleic acid chaperone activities (Hatton T, et al., (1992). J Virol 66: 5232-5241; Chu T H, et al., (2014). J Virol 88: 2530-2543), plays essential roles in viral RNA packaging and DNA synthesis, and regulates nuclear localization of HBc (Nassal M (1992). J Virol 66: 4107-4116; Yu M, Summers J (1994). J Virol 68: 4341-4348; Liao W, Ou J H (1995). J Virol 69:1025-1029.